Inhibition of proteasome activity, nuclear factor-KB translocation and cell survival by the antialcoholism drug disulfiram

Abstract

The proteasome pathway is an important target for anticancer drug development. Here, we identify the antialcoholism drug disulfiram and its analogue pyrrolidine dithiocarbamate (PDTC) as inhibitors of the 26S proteasome activity in a cell-based screening assay. As expected for proteasome inhibitors, these compounds also inhibited TNF-a-induced nuclear factor-KB (NF-KB) translocation and were cytotoxic. Disulfiram was more cytotoxic against chronic lymphocytic leukemia cells compared to peripheral blood mononuclear cells (PBMC) at clinically achievable concentrations. Proteasome and NF-KB inhibition were achieved with a potency in the same range as that of the clinically used proteasome inhibitor bortezomib. Disulfiram was also able to induce accumulation of p27Kip1 and to prolong the half-life of c-Myc, both targets for proteasome-dependent degradation. It is concluded that the previously observed antitumoral and NF-KB inhibiting activity of disulfiram and PDTC could be attributed to their inhibition of the 26S proteasome. ' 2005 Wiley-Liss, Inc.Key words: disulfiram; proteasome inhibition; nuclear factor-KB; cytotoxicityThe thiocarbamate drug disulfiram has been used extensively in the treatment of alcoholism.1 Disulfiram has also been shown to induce apoptosis in a number of tumor cell lines 2-4 as well as to reduce the growth of glioma, lung carcinoma 5 and melanoma tumors in mice. 6 However, the mechanism underlying the cytotoxic and antitumoral activity of disulfiram has not been conclusively established. It has been proposed that the induction of cell death involves generation of reactive oxygen species.5 Additionally, disulfiram has been shown to inhibit nuclear factor-KB (NF-KB) in hepatoma 4 and colorectal cancer cell lines. 2 The NF-KBinhibiting activity of the disulfiram analogue pyrrolidine dithiocarbamate (PDTC) has been attributed to its antioxidative effect.7 In this work, we propose a novel mechanism for the antitumoral and NF-KB-inhibiting activities of disulfiram. Screening of a diverse pharmacological library of compounds identified disulfiram as an inhibitor of 26S proteasome activity. Our findings suggest that the antitumor effect of disulfiram could, at least partly, be attributed to its proteasome-inhibitory action, stabilizing proteasome-degraded proteins, e.g., IKB, p27Kip1 and c-Myc, subsequently resulting in the inhibition of cell-cycle progression and induction of apoptosis. Materials and methods ChemicalsCompounds used for screening were part of the LOPAC 1280 TM library (Sigma Aldrich, St. Louis, MO). The library is a collection of pharmacologically active compounds with a diverse range of proposed target receptors and enzymes. The proteasome-inhibiting substance bortezomib was provided by Millennium Pharmaceuticals (Cambridge, MA). All other chemicals were from Sigma Aldrich. Cell linesThe stably transfected human embryo kidney cell line HEK 293 expressing the ZsProSensor-1 fusion protein (HEK 293 ZsGreen, BD Biosciences, San Jose, CA) was used for screening and evaluation of prote...

Disulfiram, a Clinically Used Anti-Alcoholism Drug and Copper-Binding Agent, Induces Apoptotic Cell Death in Breast Cancer Cultures and Xenografts via Inhibition of the Proteasome Activity

“…4). Most recently, one group reported that DSF alone at f0.16 Amol/L has proteasomeinhibitory activity under a cell-based screening assay condition (49). However, under our experimental conditions, DSF at 15 to 20 Amol/L was unable to inhibit cellular proteasome activity in cultured breast cancer cells (Figs.…”

Chemoresistance in Cancer Stem Cells and Strategies to Overcome Resistance

“…Disulfiram has been used for decades to treat alcohol abuse, and recently its potential in cancer treatment has been investigated [139]. The anti-cancer mechanisms of disulfiram are not limited to ALDH inhibition; principally, disulfiram is used to inhibit the proteasome and E3 ligases, and may also be a DNAdemethylating compound [140][141][142][143]. Disulfiram inhibits both ALDH1A1 and ALDH2 isoforms [138,144], and has shown anti-CSC effects in breast cancer and GBM [145,146].…”

Combination treatment of epilepsy with ketogenic diet and concurrent pharmacological inhibition of cytochrome P450 2E1

“…Disulfiram was first characterized as an inhibitor of aldehyde dehydrogenase, and as such has been used for more than fifty years to prevent relapse in alcohol addicts. Subsequently, disulfiram was found to also inhibit dopamine-β-hydroxylase [43], CYP2E1 [44], and proteasomes [45]; novel applications based on these effects have been proposed [46]. The drug is metabolized rapidly, and it is the metabolites that are responsible for the various inhibitory effects.…”